FANCF基因沉默增强多药耐药多发性骨髓瘤细胞株RPMI8226/R对马法兰的敏感性  

Knockdown of FANCF gene potentiates the sensitivity to melphalan in multidrug resistant multiple myeloma RPMI8226 / R cells

作　　者：裴梦婉 靳鸣 陈小琼[1] 肖晖[1] 

机构地区：[1]武汉大学中南医院血液内科,湖北武汉430071

出　　处：《现代肿瘤医学》2016年第18期2846-2850,共5页Journal of Modern Oncology

摘　　要：目的:使用小片段干扰RNA(siRNA)沉默FANCF基因表达,观察其对FA/BRCA途径的影响及对多药耐药多发性骨髓瘤细胞株RPMI8226/R对交联剂马法兰的敏感性变化。方法:FANCF siRNA瞬时转染多药耐药细胞株RPMI8226/R,采用CCK-8法检测转染前后细胞对马法兰的敏感性变化,RT-PCR,westernblot法检测FA/BRCA途径中FANCF、FANCD2、BRCA2基因表达量的改变,彗星实验检测DNA损伤,流式细胞仪检测细胞凋亡率。结果:FANCF siRNA转染耐药骨髓瘤细胞后,对马法兰的IC50值由17.29μmol/L降至4.88μmol/L;western-blot检测出瞬时转染FANCF siRNA后FA/BRCA途径中FANCF、FANCD2、BRCA2蛋白表达量明显较转染前减低;彗星实验检测出转染后耐药细胞的DNA损伤增多;转染后马法兰诱导细胞凋亡率由(77.67±0.62)%升高至(88.37±0.25)%,提示FANCF基因沉默可使耐药RPMI8226/R细胞对化疗药物马法兰重新恢复敏感性。结论:FANCF基因沉默可抑制FA/BRCA途径中FANCD2及下游BRCA2基因表达,从而阻断FA/BRCA途径介导的DNA修复,增强马法兰对多药耐药骨髓瘤细胞株RPMI8226/R的细胞毒性,增加细胞凋亡率,为解决临床耐药问题提供一个新的思路。Objective： To investigate the change of sensitivity to melphalan and the relationship with FA/BRCA pathway when FANCF gene is knocked down by small interfering RNA（ siRNA） in multidrug resistant human multiple myeloma cell line RPMI8226 / R. Methods： Small interfering RNA（ siRNA） targeting FANCF gene was used to transfect RPMI8226 / R cells. Then we used a series of tests to detect the drug sensitivity to melphalan,gene expression in FA / BRCA pathway,and DNA damage caused by melphalan pre- and post- transfection in RPMI8226 / R cells.Results： IC50 value to melphalan decreased from 17. 29μmol / L to 4. 88μmol / L pre- and post- transfection,suggesting that depletion of FANCF restored the drug- resistant cell line RPMI8226 / R sensitivity to melphalan. The proteins associated with FA / BRCA pathway,FANCF,FANCD2,BRCA2 obviously decreased after treatment with FANCF siRNA. Melphalan- induced DNA damage and apoptosis percentages increased from 77. 67% to 88. 37% in FANCF silencing cells after treatment with siRNA. Conclusion： Downregulating expression of FANCF gene by siRNA could potentiate sensitivity to melphalan,inhibit the DNA interstrand crosslink repair,and enhance the DNA damage and apoptosis in melphalan- resistance multiple myeloma cells by blocking the FA / BRCA pathway in RPMI8226 / R cells.

关 键 词：FANCF FA/BRCA途径 多药耐药 RNA干扰 马法兰 FA核心蛋白 

分 类 号：R733.2[医药卫生—肿瘤]